TTV基因分型的研究

目的调查华南地区 ＴＴＶ感染流行情况和基因分型。方法选取 ＴＴＶ开放读码枢１的保守序列作内外引物, Ｇ１、 Ｇ２、 Ｇ３、 Ｇ４、 Ｇ５、 Ｇ６同源性极高的系列 ２ １６０— ２ １９６ ｎｔ作包被探针,异源性大于 ２０％的序列作显色探针Ｇ１、Ｇ２、Ｇ３、Ｇ４、Ｇ５、Ｇ６,采用ＰＣＲ微板核酸杂交ＥＬＩＳＡ技术对３８０例肝功能损害的肝炎患者血清进行ＴＴＶ检测及ＴＴＶ基因分型研究。结果检测出６１例ＴＴＶ阳性患者,检出率为１６．０％。对６１例ＴＴＶ进行基因分型,Ｇ１型４４例,Ｇ２型５例,Ｇ１、Ｇ２混合型感染１０例,Ｇ３、Ｇ４各１例,尚未发现Ｇ５、Ｇ６型。慢性肝炎中ＴＴＶ感染者与急性肝炎中ＴＴＶ感染者比例两者相近。结论在华南地区的肝炎疾病中, ＴＴＶ有较高的感染率; ＴＴＶ存在四个基因型,主要为Ｇ１型,其次为Ｇ２型,尚未发现Ｇ５、Ｇ６型。     

延边地区45例乙型与丙型肝炎患者合并TTV感染的研究

观察延边地区慢性乙肝和慢性丙肝患者ＴＴＶ感染状况，并研究其基因型。对４５例慢性病毒性肝炎患者，通过ＰＣＲ检测其ＴＴＶ，并对ＴＴＶ进行核苷酸序列分析。慢性肝炎病人确实有ＴＴＶ感染，ＴＴＶ亚型可分为Ⅰ型和Ⅱ型，Ⅱ型与日本东京的亚型一致。     

原位杂交检测肝组织中TTV DNA的研究

研究ＴＴＶＤＮＡ在肝炎患者肝组织中的表达 ,证实ＴＴＶ是一种新型肝炎病毒。用地高辛标记ＴＴＶＤＮＡ探针 ,以原位杂交技术对血清ＰＣＲ检测为单一ＴＴＶ感染者 13例 ,ＴＴＶ、ＨＢＶ混合感染者 4例和非甲～庚型、非ＴＴＶ感染的肝炎患者 6例的肝组织进行检测 ,同时进行肝组织学、免疫组织化学、肝功能和临床特点等分析。结果表明 ,2 3例肝组织中ＴＴＶＤＮＡ阳性 15例 ( 65.2 % ) ,其中单一ＴＴＶ感染者11例 ( 84 .6% ) ,ＴＴＶ、ＨＢＶ混合感染者 3例 ( 75.0 % ) ,非甲～庚型、非ＴＴＶ型肝炎 1例 ( 16.7% )。按肝炎临床分型 ,急性肝炎…     

TTV与其它肝炎病混合感染及其基因型的研究

研究TTV与其它肝炎病混合感染对肝脏病变的影响及本地区TTVDNA的基因型。选TTV ORF1的保守序列作内外引物,采用微板核酸杂交－ELISA方法检测患者血清TTVDNA并对检测结果和临床资料进行统计学分析。选取异源性大于50％的序列作显色探针Ⅰ和显色探针Ⅱ,进行分型研究。学生、非甲－非戊型肝炎、慢性乙型肝炎和肝硬化患者血清TTV阳性率分别为3．3％、14．3％、12％、16％。TTV阳性和TTV阴性的慢性乙型肝炎及肝硬化患者,在年龄、性别、ALT和TBil之间无显著差异（P＞0．05）。本地区流行的TTV可分为两个主要的基因型,即I型（66．75）、Ⅱ型（25％）,部分存在混合感染（8．3％）。TTV可能与HBV、HCV有类似的传播途径,故常重叠感染。TTV与乙型肝炎及肝硬化混合感染后不影响两者的肝脏病变。TTV不是本地区非甲－非戊型肝炎的主要病因。     

非甲－非庚型肝炎肝组织中经输血传播病毒DNA的原位检测

目的探讨非甲－非度型病毒性肝炎患者肝组织中经输血传播病毒（ｔｒａｎｓｆｕｓｉｏｎｔｒａｎｓｍｉｔｔｅｄｖｉｒｕｓ,ＴＴＶ）ＤＮＡ的表达。方法采用地高辛素标记ＴＴＶＤＮＡ探针以原位杂交技术对１９例血清学甲、戊型肝炎病毒标志物、免疫组织化学检测肝组织中ＨＧＶＮＳＳ均用性的急、慢性病毒性肝炎患者肝组织进行检测。结果１９例肝组织中６例检出ＴＴＶ基因阳性（３１．５８％）,其中急性轻型肝炎４例（３０．８０％）,慢性肝炎２例（３３．３３％）。ＴＴＶＤＮＡ表达于肝细胞核或胞浆内。在急性肝炎,ＴＴＶ阳性细胞弥漫分布于肝小叶内,慢性肝炎于汇管区附近较为密集。６例中５例患者有转氨酶增高。结论非甲－非庚型病毒性肝炎肝组织中ＴＴＶＤＮＡ的检出表明ＴＴＶ为一种新型的嗜肝性病毒。     

经输血传播病毒(TTV)在血液透析患者中感染状况及基因分析

目的 研究我国血液透析患者人群中经输血传播病毒（ＴＴＶ）的感染及基因变异状况。方法 设计特异性引物采用半巢式ＰＣＲ方法对北京市２所综合性大医院５０例血液透析患者血清标本进行ＴＴＶ ＤＮＡ的检测并将产物克隆及序列分析。结果 经过半巢式ＰＣＲ检测，血透患者ＴＴＶ ＤＮＡ阳性率为１４％。对其中４例标本中分离出的ＴＴＶ ２７０个碱基核酸片段序列分析表明：基因同源性为９９．１％￣１００％，推测的编码氨基酸序     

非甲—庚型肝炎患者TT病毒感染的检测及序列分析

在急慢性输血后肝炎、散发性肝炎及爆发性肝炎中，有相当比例的患者用现有甲、乙、丙、丁、戊、庚型肝炎实验室诊断方法尚不能分型，在排除已知的致病因素外，仍有３．５０～１５％的肝炎病人病因不明。通常将这些不明原因的肝炎称之为非甲－庚型肝炎。１９９７年日本学者报道发现一种新的肝炎病毒，并命名为ＴＴ病毒（ＴＴＶ）［１］。我们根据日本报道的ＴＴＶ基因序列，建立套式聚合酶链反应（ｎｅｓｔｅｄＰＣＲ）方法，在我国北京地区非甲庚型肝炎患者中检测出ＴＴＶ基因，并进行了序列分析，现将结果报道如下。材料与方法一、标…     

人巨细胞病毒糖蛋白B基因的分型

目的建立人巨细胞病毒(HCMV)糖蛋白B(gB)基因的套式聚合酶链反应(nPCR)加限制性长度多态性分析(RFLP)的分型方法。方法34例HCMV感染患儿的11份全血标本和23份尿标本，用nPCR扩增gB基因片段，RFLP对gB基因进行分型，并通过测序进行验证。结果34例HCMV感染患儿中，检出gBⅠ型13例，gBⅡ型12例，gBⅢ型9例，未发现gBⅣ型。测序结果与基因库中相应的标准株序列进行比较，Ⅰ、Ⅱ和Ⅲ型的同源性分别为98．1％～99．6％、98．9％～100％、97．3％～98．9％。结论nPCR检测HCMV感染敏感特异，RFLP对HCMV的gB基因分型明确可靠。     

新疆地区输血传播病毒的检测

随着检测技术的发展，对肝炎病毒的认识更加深刻。除甲～戊型肝炎和已知的其它致病因素外，仍有１０％～２０％的因输血或散发性肝炎不能用现有的检测方法检出其病毒标志。１９９７年Ｎｉｓｈｉｚａｗａ等报告了一种新发现的，与人类肝炎相关的ＤＮＡ病毒ＴＴＶ。为了解新疆地区ＴＴＶ感染情况，我们用日本学者报道的ＴＴＶ核苷酸序列设计了两对引物，检测了新疆地区的１２８例各种肝炎患者和４１例血液透析患者血清，证实新疆地区存在ＴＴＶ感染的流行，现将检测结果和部分ＴＴＶ阳性核苷酸测序结果总结报道如下。材料与方法一、病例来源本…     

斑点杂交法检测肝病患者TTV感染状况初探

目的 探讨斑点杂交法在检测肝病患者ＴＴ病毒（ＴＴＶ）感染状况中的应用价值。方法 提抽患者血清中ＤＮＡ,分别采用聚合酶链反应（ＰＣＲ）及斑点杂交技术检测其中的ＴＴＶ病毒ＤＮＡ。结果 在３０份非甲～非戊型肝炎患者血清中,ＰＣＲ方法检出１１份为ＴＴＶ阳性,检出率为３６．７％。采用斑点杂交法检出３份阳性血清,阳性率为１０％。制备的探针具有ＴＴＶ特异性,可检出相当于１０＾６拷贝的病毒ＤＮＡ。结论 以地高辛标     

TT virus infection in Italy: prevalence and genotypes in healthy subjects, viral liver diseases and asymptomatic infections by parenterally transmitted viruses

This study was aimed to evaluate TT virus prevalence in subjects with hepatitis B virus (HBV), hepatitis C virus (HCV) and human immunodeficiency virus (HIV) infections in patients affected by hepatitis of unknown origin (non-A–non-E hepatitis) and in healthy subjects who had not been exposed to HBV, HCV and HIV. A total of 317 subjects were tested; 40 were HBsAg asymptomatic carriers, 57 subjects were anti-HCV positive (45 without chronic hepatitis and 12 with HCV-related chronic hepatitis), and 27 had chronic non-A–non-E hepatitis. Fifty-seven subjects were intravenous drug users (IVDUs) (52 with HCV or/and HIV infections), seven patients underwent a liver transplant for fulminant hepatitis and 137 were healthy subjects from the general population. Overall, TTV-DNA was detected in 62 subjects (19.6%): in 17.9% of the HBsAg carriers, in 14% of the anti-HCV-positive patients (in 8.3% and in 15.5% of patients with and without chronic hepatitis, respectively), in 22.2% of non-A–non-E hepatitis patients, in 22.8% of IVDUs, in 57.1% of fulminant hepatitis patients. TTV-DNA was also found in 20.4% healthy subjects. The prevalence in the different subgroups was not statistically different. The genotypes were identified in 40 of the 62 (64.5%) TTV-DNA positive samples: genotype 1a in 17.5%, 1b in 27.5%, genotype 2 in 27.5%, genotype 3 in 15.0%, genotype 4 in 5.0% and genotype 5 in 7.5%; the genotype distribution in the subsets of patients was not significantly different. In conclusion, this study showed that TTV infection is common in Italy; it is widespread throughout the entire population and five genotypes are present in Sardinia. Our results further dismiss the role of TTV as cofactor in influencing the clinical course of infections with other hepatitis viruses as well as the role of HIV in enhancing TTV transmission and replication.     

Transfusion-transmitted virus infection in China: Prevalence in blood donors and in patients with liver diseases

BACKGROUND: Prevalence of transfusion-transmitted virus (TTV) infection among blood donors and in patients with liver diseases in China was studied. METHODS: DNA was extracted from serum and amplified by seminested polymerase chain reaction with reported primer sets from a conserved region of the TTV genome. RESULTS: TT Virus DNA was detected in 55 of 196 blood donors (28%); 31% (40 of 127) in the north and 22% (15 of 69) in the south. TT Virus DNA was also detected in 14 of 31 patients (45%) with non-A-non-G fulminant hepatitis and in eight of 25 patients (32%) with non-A-non-G chronic hepatitis. The rate of TTV viraemia in these patients with liver disease was comparable to that in blood donors. TT Virus DNA sequencing of 12 isolates showed that the prevalence of genotype 2 was significantly higher than that reported in Japan (66.7 vs 2.6%, P < 0.001). Furthermore, genotyping assays based on restriction fragment length polymorphism were carried out on all 88 TTV DNA-positive samples. It was found that 42 isolates (47.7%) belonged to genotype 1 and 40 (45.5%) to genotype 2. It was of particular interest that the prevalence of genotype 1 in patients with non-A-non-G fulminant hepatitis was significantly higher than that in blood donors (10/14 vs 22/55, P < 0.05). CONCLUSIONS: The data indicate that TTV infection is common in China and that the pathogenic potential of TTV toward the liver (if any) may differ between genotypes.     

TT virus infection in adult beta-thalassemia major patients

BACKGROUND AND OBJECTIVES: Patients with thalassemia-major are at risk of blood-borne viral infections. TT virus (TTV), a single stranded, circular DNA virus, has recently been found to be associated with acute and chronic hepatitis. The aims of this study were to assess the prevalence of TTV infection in adult patients with transfusion-dependent thalassemia, and to evaluate the clinical significance of TTV. DESIGN AND METHODS: We studied 68 adult patients with thalassemia major, 97% of whom were hepatitis C virus (HCV) antibody positive. TTV DNA was amplified from serum by heminested polymerase chain reaction (PCR). Direct sequencing of PCR products was used to establish TTV genotypes. RESULTS: TTV DNA was detected in 47 patients (69.1%). Sequence analysis of PCR products identified TTV genotype 1b as the most common viral genotype in this group. Patients co-infected by HCV and TTV had a significantly higher histologic grade score than patients with isolated HCV infection (5.1+/-2.7 vs. 2.8+/-1.7, p=0.02) while the stage score was not significantly different. INTERPRETATION AND CONCLUSIONS: TTV is highly prevalent among Italian multiply transfused patients. In these patients TTV viremia appears to affect the necro-inflammatory activity of hepatitis C, though no evidence of an effect on the evolution of fibrosis was found.     

Clinical characteristics and distribution of genotypes of TT virus infection in a hepatitis C virus-hyperendemic township of a hepatitis B virus-endemic country (Taiwan)

BACKGROUND: The prevalence of TT virus (TTV) viremia, without definite clinical significance, has been reported to be higher among chronic hepatitis C patients. The status and clinical characteristics of TT virus (TTV) infection and distribution of TTV genotypes in a hepatitis C virus (HCV) hyperendemic township (Masago community) in a hepatitis B virus (HBV) endemic country (Taiwan) were investigated. METHODS: Sera from 100 Masago residents were tested for alanine aminotransferase (ALT) and markers of HBV, HCV and GB virus C/hepatitis G virus (GBV-C/HGV) and TTV-DNA. Sera of 250 blood donors as a control group were tested for TTV-DNA. Sera of Masago residents and blood donors with positive TTV-DNA were directly sequenced, and phylogenetic analyses were performed subsequently. RESULTS: The prevalences of TTV viremia in different age groups among individuals from Masago were significantly higher than that among blood donors. In regard to the subtypes of TTV, 23, seven, two, eight, one, six and one isolate were related to the genotypes 1a, 1b, 2a, 2b, 3, 4 and 5, respectively, from Masago and 21, 14, one, nine and three isolates were related to the genotypes 1a, 1b, 2a, 2b, and 4, respectively, from donors. No clinical or virological factor was associated with TTV viremia or TTV genotypes. CONCLUSIONS: TT Virus prevalence was higher among HCV hyperendemic township residents than blood donors with similar genotype distributions (genotype 1 was the most prevalent) in Taiwan. Neither TTV viremia nor a particular genotype was associated with HBV, HCV or GBV-C/HGV infection and abnormal ALT levels.     

长春地区TT病毒的基因分型及其临床意义

为调查各种急、慢性肝炎中TT病毒的感染状况及临床意义,并检测TTV基因的分型。利用半套式PCR（semi-nested PCR）方法检测了TTV-DNA。利用邻近丁（neighbor-joining）法画出系统树。TTV-DNA的阳性率在非甲非乙非丙型急性肝炎中为42.3%,在非乙非丙型慢性肝炎中为45.5%。基因型可分为1a、1b、2a、2b等型。TTV-DNA在非甲非乙非丙型急性非乙非丙型慢性肝炎中的感染率最高;在TT病毒与乙型、丙型肝炎病毒混合感染的慢性肝炎中,TT病毒干涉乙型及丙型肝炎病毒造成的肝细胞损伤的可能性很小。     

Impact of TT virus infection in acute and chronic, viral- and non viral-related liver diseases

BACKGROUND/AIMS: The prevalence and pathogenicity of TT virus, recently identified in patients with non A-non G post-transfusional hepatitis, are questioned. METHODS: We investigated the impact of this new viral infection in a large series of patients with non A-non G, cryptogenic, non-viral and viral-related, acute and chronic liver diseases (n=577) and blood donors (n=300). TTV DNA was detected in serum by hemi-nested polymerase chain reaction. Phylogenetic analysis was performed in 13 isolates. RESULTS: TTV DNA was detected in 6/25 and 15/127 patients with cryptogenic non A-non G acute and chronic liver disease, respectively. TTV DNA positive subjects with post-transfusional acute hepatitis scored negative before transfusion. TTV prevalence was increased in patients with cryptogenic non A-non G acute and chronic liver disease compared to blood donors (6/300; p<0.001) and non-viral-related chronic liver diseases (6/137; p<0.05). TTV/HBV coinfection was frequently identified (35/147), but this was not the case for HCV-infected subjects (4/77). Transaminase activity or liver histological score was not significantly increased among TTV positive, HBV infected or non A-non G patients. The HBV infection and Mediterranean origin were the risk factors associated with TTV infection. The majority of analysed sequences clustered in genotype 1 (8=1b; 3=1a). Two isolates showed homology to genotype 2. CONCLUSIONS: These results support the view that TTV is a widely spread infectious agent with a weak pathogenicity. It raises the possibility, however, that TTV might be implicated in a few cases of acute and chronic non A-non G hepatitis. TTV-DNA-analysed sequences are related to genotypes 1 and 2 described in Europe.     

Clinical impact of genotype 1 TT virus infection in patients with chronic hepatitis C and response of TT virus to alpha-interferon

BACKGROUND: The relationship between genotype 1 TT virus (TTV) infection and the status of chronic hepatitis C was studied. METHODS: A total of 52 patients with chronic hepatitis C who were treated with interferon (IFN)-alpha were enrolled in the present study. Of those, 12 were infected with genotype 1 TTV and 40 were uninfected. RESULTS: Clinical backgrounds, including mean age, sex, blood transfusion history, serum alanine aminotransferase (ALT) level, and the results of liver biopsy did not differ between patients with and without genotype 1 TTV infection. The distribution of hepatitis C virus (HCV) genotypes did not differ between the two groups of patients, but TTV-infected patients tended to have a lower serum HCV-RNA level than uninfected patients (median (range) 26.0 (< 1-460) vs 135 (1.2-740) kilo copies/mL, respectively; P = 0.065). Patients with a sustained response of HCV to IFN-alpha were significantly more common in TTV-infected than -uninfected patients (58 vs 23%, respectively; P = 0.018). Multivariate logistic regression analysis revealed that patients with a sustained response of HCV correlated significantly with the serum HCV-RNA level (P = 0.006), but not with the presence or absence of genotype 1 TTV infection (P = 0.161). Serum TTV-DNA decreased with IFN-alpha therapy in all 12 patients and remained negative in six patients even after treatment. There was no correlation between patients with a sustained response of HCV and the same of TTV. Serum ALT levels correlated with changes in the status of HCV viremia, but not with changes in the status of TTV viremia. CONCLUSIONS: An opposing relationship between HCV and TTV proliferation was suggested, but coinfection with genotype 1 TTV did not affect the status of chronic hepatitis C.     

TT virus infection among blood donors and patients with non-B, non-C liver diseases in Korea

BACKGROUND/AIMS: A novel virus, designated the TT virus (TTV), was isolated from the serum of a patient with posttransfusion hepatitis of unknown etiology, in Japan. Subsequently, TTV was suggested to be a causative agent in a proportion of cases with cryptogenic hepatitis in Japan. This study aimed to elucidate the significance of TTV infection in cases with cryptogenic liver disease in Korea, a neighbor of Japan. METHODS: The prevalence of TTV infection was studied in 120 patients with liver diseases, including 85 patients diagnosed as having non-B, non-C liver diseases. As controls, 220 blood donors were also examined. TTV DNA was detected by polymerase chain reaction, and the sequence was analyzed by phylogenetic analysis. RESULTS: Fourteen (14.0%) of 100 accepted blood donors, 23 (19.2%) of 120 rejected blood donors, and 15 (17.6%) of 85 patients with non-B, non-C liver diseases were positive for TTV DNA. The prevalences of TTV infection among these groups were not significantly different. Phylogenetic analysis suggested the existence of four major genotypes of TTV The proportions of each genotype among patients with non-B, non-C liver diseases were not different from those among accepted blood donors. CONCLUSIONS: TTV exists in Korea, but the prevalence among patients with non-B, non-C liver diseases was almost the same as that among blood donors. TTV may not be the main causative agent of cryptogenic liver disease in Korea. The relationship between non-B, non-C liver diseases and TTV genotype remains unclear, although TTV can be classified into four genotypes.     

Prevalence and Genotypic Variability of TTV in HIV-Infected Patients

TT virus is a small, circular DNA virus, that has been associated with transfusion hepatitis. We sought to determine the prevalence of TT virus (TTV) in patients with human immunodeficiency virus (HIV) infection and to characterize the virus in terms of genotypic variability and in the relationship to CD4⁺, HIV viral loads, HCV/HIV coinfection, and ALT abnormalities. A cross-sectional analysis of HIV-infected patients in the United States, including 86 HIV-positive subjects and 118 HIV-negative controls was performed. TTV was detected using a seminested PCR technique. Samples underwent cloning and sequence analysis and/or RFLP to determine genotype. Thirty-eight percent of HIV-positive patients had TTV infection versus 14.4% of patients within the matching cohort (P = 0.0009). The highest rate of TTV infection was in patients with concurrent HCV/HIV infection (54% vs 30%, P = 0.038). HIV-infected subjects with TTV had lower ALT levels than those without TTV (P = 0.036). Intravenous drug use was the leading factor associated with TTV positivity among HIV-positive subjects. Mixed genotypes were more common in those with HIV. Therefore, TTV prevalence, ALT levels, and genomic heterogeneity of TTV all seem to be altered in patients with HIV.